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Coexpression of redox partners increases the hydrocortisone (cortisol) production efficiency in CYP11B1 expressing fission yeast Schizosaccharomyces pombe

Paper ID Volume ID Publish Year Pages File Format Full-Text
24811 43538 2008 9 PDF Available
Title
Coexpression of redox partners increases the hydrocortisone (cortisol) production efficiency in CYP11B1 expressing fission yeast Schizosaccharomyces pombe
Abstract

Cytochromes P450 play a vital role in the steroid biosynthesis pathway of the adrenal gland. An example of an essential P450 cytochrome is the steroid 11β-hydroxylase CYP11B1, which catalyses the conversion of 11-deoxycorticol to hydrocortisone. However, despite its high biotechnological potential, this enzyme has so far been unsuccessfully employed in present-day biotechnology due to a poor expression yield and inherent protein instability. In this study, CYP11B1 was biotransformed into various strains of the yeast Schizosaccharomyces pombe, all of which also expressed the electron transfer proteins adrenodoxin and/or adrenodoxin reductase – central components of the mitochondrial P450 system – in order to maximise hydrocortisone production efficiency in our proposed model system. Site-directed mutagenesis of CYP11B1 at positions 52 and 78 was performed in order to evaluate the impact of altering the amino acids at these sites. It was found that the presence of an isoleucine at position 78 conferred the highest 11β-hydroxylation activity of CYP11B1. Coexpression of adrenodoxin and adrenodoxin reductase appeared to further increase the 11β-hydroxylase activity of the enzyme (3.4 fold). Adrenodoxin mutants which were found to significantly enhance enzyme efficiency in other cytochromes in previous studies were also tested in our system. It was found that, in this case, the wild type adrenodoxin was more efficient. The new fission yeast strain TH75 coexpressing the wild type Adx and AdR displays high hydrocortisone production efficiency at an average of 1 mM hydrocortisone over a period of 72 h, the highest value published to date for this biotransformation. Finally, our research shows that pTH2 is an ideal plasmid for the coexpression of the mitochondrial electron transfer counterparts, adrenodoxin and adrenodoxin reductase, in Schizosaccharomyces pombe, and so could serve as a convenient tool for future biotechnological applications.

Keywords
Adrenodoxin; Adrenodoxin reductase; Hydrocortisone; CYP11B1; Fission yeast; Schizosaccharomyces pombe; Steroid 11β-hydroxylation
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Coexpression of redox partners increases the hydrocortisone (cortisol) production efficiency in CYP11B1 expressing fission yeast Schizosaccharomyces pombe
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 133, Issue 3, 1 February 2008, Pages 351–359
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us